LOCUS       sequence-417168-        5715 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of ATG13 (460-517) mutant.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5715)
  AUTHORS   Chen M, Ren X, Nguyen TN, Khuu G, Cook ASI, Lazarou M, Hurley JH
  TITLE     Structure and activation of the human autophagy-initiating 
            ULK1C:PI3KC3-C1 supercomplex
  JOURNAL   bioRxiv 2023.06.01.543278
REFERENCE   2  (bases 1 to 5715)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            2023.06.01.543278"
COMMENT     Sequence Label: pCAG-GST-ATG13 (460-517)-F466D/F470D
FEATURES             Location/Qualifiers
     source          1..5715
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        385..661
                     /label=chicken beta-actin promoter
     intron          662..1678
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1602..1624
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1686..1705
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1753..1762
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1759..2415
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MASPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNK
                     KFELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIR
                     YGVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVV
                     LYMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     2373..2395
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             2443..2463
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(2736..2755)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2801..2856
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2855..2874)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3217..3233)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3217..3233)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3230..3252)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3241..3257
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(3265..3271,3272..3289,3290..3295))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3310..3331
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3390..3585
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3436..3571
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3591..3725
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3628..3647)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3682..3701
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3793..3810)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3964..4552)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4044..4063)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4723..5514,5515..5583))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5346..5365
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5584..5688)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
       61 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      121 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      181 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
      241 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      301 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      361 tattagtcat cgctattacc atggtcgagg tgagccccac gttctgcttc actctcccca
      421 tctccccccc ctccccaccc ccaattttgt atttatttat tttttaatta ttttgtgcag
      481 cgatgggggc gggggggggg gggggcgcgc gccaggcggg gcggggcggg gcgaggggcg
      541 gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgcgctgcc ttcgccccgt gccccgctcc gccgccgcct cgcgccgccc
      721 gccccggctc tgactgaccg cgttactccc acaggtgagc gggcgggacg gcccttctcc
      781 tccgggctgt aattagcgct tggtttaatg acggcttgtt tcttttctgt ggctgcgtga
      841 aagccttgag gggctccggg agggcccttt gtgcgggggg agcggctcgg ggggtgcgtg
      901 cgtgtgtgtg tgcgtgggga gcgccgcgtg cggctccgcg ctgcccggcg gctgtgagcg
      961 ctgcgggcgc ggcgcggggc tttgtgcgct ccgcagtgtg cgcgagggga gcgcggccgg
     1021 gggcggtgcc ccgcggtgcg gggggggctg cgaggggaac aaaggctgcg tgcggggtgt
     1081 gtgcgtgggg gggtgagcag ggggtgtggg cgcgtcggtc gggctgcaac cccccctgca
     1141 cccccctccc cgagttgctg agcacggccc ggcttcgggt gcggggctcc gtacggggcg
     1201 tggcgcgggg ctcgccgtgc cgggcggggg gtggcggcag gtgggggtgc cgggcggggc
     1261 ggggccgcct cgggccgggg ggggctcggg ggaggggcgc ggcggccccc ggagcgccgg
     1321 cggctgtcga ggcgcggcga gccgcagcca ttgcctttta tggtaatcgt gcgagagggc
     1381 gcagggactt cctttgtccc aaatctgtgc ggagccgaaa tctgggaggc gccgccgcac
     1441 cccctctagc gggcgcgggg cgaagcggtg cggcgccggc aggaaggaaa tgggcgggga
     1501 gggccttcgt gcgtcgccgc gccgccgtcc ccttctccct ctccagcctc ggggctgtcc
     1561 gcggggggac ggctgccttc gggggggacg gggcagggcg gggttcggct tctggcgtgt
     1621 gaccggcggc tctagagcct ctgctaacca tgttcatgcc ttcttctttt tcctacagct
     1681 cctgggcaac gtgctggtta ttgtgctgtc tcatcatttt ggcaaagaat tcctcgatcg
     1741 acggtatcga tagccaccat ggcatcccct atactaggtt attggaaaat taagggcctt
     1801 gtgcaaccca ctcgacttct tttggaatat cttgaagaaa aatatgaaga gcatttgtat
     1861 gagcgcgatg aaggtgataa atggcgaaac aaaaagtttg aattgggttt ggagtttccc
     1921 aatcttcctt attatattga tggtgatgtt aaattaacac agtctatggc catcatacgt
     1981 tatatagctg acaagcacaa catgttgggt ggttgtccaa aagagcgtgc agagatttca
     2041 atgcttgaag gagcggtttt ggatattaga tacggtgttt cgagaattgc atatagtaaa
     2101 gactttgaaa ctctcaaagt tgattttctt agcaagctac ctgaaatgct gaaaatgttc
     2161 gaagatcgtt tatgtcataa aacatattta aatggtgatc atgtaaccca tcctgacttc
     2221 atgttgtatg acgctcttga tgttgtttta tacatggacc caatgtgcct ggatgcgttc
     2281 ccaaaattag tttgttttaa aaaacgtatt gaagctatcc cacaaattga taagtacttg
     2341 aaatccagca agtatatagc atggcctttg cagggctggc aagccacgtt tggtggtggc
     2401 gaccatcctc caaaatcgga tgattacgat atcccaacga ccgaaaacct gtattttcag
     2461 ggtggtaccc ctatggatct gggaaccgac tacagggagg atcagaatcc ccctcagctc
     2521 agtagcctga gcattgacat cggtgcccag tccatggctg aggacctcga ttctctgcct
     2581 gaaaaactgg ctgtgcacga gaagaacgtc agagaattcg acgcatttgt ggagaccctg
     2641 cagtgactcg aggaattcac tcctcaggtg caggctgcct atcagaaggt ggtggctggt
     2701 gtggccaatg ccctggctca caaataccac tgagatcttt ttccctctgc caaaaattat
     2761 ggggacatca tgaagcccct tgagcatctg acttctggct aataaaggaa atttattttc
     2821 attgcaatag tgtgttggaa ttttttgtgt ctctcactcg gaaggacata tgggagggca
     2881 aatcatttaa aacatcagaa tgagtatttg gtttagagtt tggcaacata tgcccatatg
     2941 ctggctgcca tgaacaaagg ttggctataa agaggtcatc agtatatgaa acagccccct
     3001 gctgtccatt ccttattcca tagaaaagcc ttgacttgag gttagatttt ttttatattt
     3061 tgttttgtgt tatttttttc tttaacatcc ctaaaatttt ccttacatgt tttactagcc
     3121 agatttttcc tcctctcctg actactccca gtcatagctg tccctcttct cttatggaga
     3181 tccctcgacc tgcagcccaa gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
     3241 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     3301 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     3361 gtcgggaaac ctgtcgtgcc agcggatccg catctcaatt agtcagcaac catagtcccg
     3421 cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat
     3481 ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc
     3541 cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct aacttgttta
     3601 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     3661 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     3721 ggatccgctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     3781 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     3841 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     3901 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     3961 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     4021 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     4081 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     4141 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     4201 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     4261 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     4321 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     4381 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     4441 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     4501 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     4561 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     4621 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     4681 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     4741 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     4801 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     4861 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     4921 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     4981 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     5041 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     5101 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     5161 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     5221 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     5281 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     5341 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     5401 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     5461 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     5521 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     5581 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     5641 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     5701 aaaagtgcca cctgg
//
